Team:HKUST/Notebook

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        <li><a href="http://2010.igem.org/Team:HKUST">HOME</a></li>
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        <li><a href="http://2010.igem.org/Team:HKUST/Team">Here we are!</a></li>
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        <li><a>Project</a><li>
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        <li class="project"><a href="http://2010.igem.org/Team:HKUST/Project">Abstract</a></li>
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<li class="project"><a href="http://2010.igem.org/Team:HKUST/Project/Background">Background</a></li>
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        <li class="project"><a href="http://2010.igem.org/Team:HKUST/Project/Experiment Design">Experiment Design</a></li>
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        <li class="project"><a href="http://2010.igem.org/Team:HKUST/Project/Results and Discussion">Results and Discussion</a></li>
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        <li class="project"><a href="http://2010.igem.org/Team:HKUST/Project/Materials and Methods">Materials and Methods</a></li>
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        <li><a href="http://2010.igem.org/Team:HKUST/biobrick">Biobricks</a></li> 
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        <li><a href="http://2010.igem.org/Team:HKUST/Human_Practice">Human Pratice</a></li>   
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<li><a href="http://2010.igem.org/Team:HKUST/Notebook">Lab Notebook</a></h3></li>
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        <li><a href="http://2010.igem.org/Team:HKUST/Acknowledgement">Acknowledgement</a></li>
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        <li><a href="http://2010.igem.org/Team:HKUST/Safety">Biosafety</a></li>
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<div id="leftpart"> <h1>Lab Notebook</h1></div>
<div id="leftpart"> <h1>Lab Notebook</h1></div>
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<div id="leftpart">
<div id="leftpart">
<div id="date">25/06/2010 </div>
<div id="date">25/06/2010 </div>
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<div id="logbook">PMH4 containing mCherry and PBI121 conatining gusA reporter gene were successfully transferred into <em>E.coli</em>  DH10B competent cells. </div>
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<div id="logbook">Plasmid PMH4 containing mCherry and plasmid PBI121 conatining gusA reporter gene were successfully transferred into <em>E.coli</em>  DH10B competent cells. </div>
</div>
</div>
<div id="leftpart">
<div id="leftpart">
<div id="date">12/07/2010 </div>
<div id="date">12/07/2010 </div>
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<div id="logbook">Fusion PCR of agrC-mCherry  was performed and products were confirmed by agarose gel electrophoresis.</div>
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<div id="logbook">Fusion PCR of <em>agrC-mCherry</em> was performed and products were confirmed by agarose gel electrophoresis.</div>
</div>
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<div id="date">22/07/2010 </div>
<div id="date">22/07/2010 </div>
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<div id="logbook"><em>E.coli</em> DH10B competent cells were successfully transformed with pMG36e.</div>
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<div id="logbook">Transform <em> E.coli</em> DH10B competent with plasmid pMG36e.</div>
</div>
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<div id="leftpart">
<div id="leftpart">
<div id="date">28/07/2010 </div>
<div id="date">28/07/2010 </div>
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<div id="logbook">Enzyme digested the vector  and confirmed plasmid pMG36e.</div>
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<div id="logbook">Plasmid pMG36e was enzyme digested and confirmed..</div>
</div>
</div>
   
   
<div id="leftpart1">
<div id="leftpart1">
<div id="date">30/07/2010 </div>
<div id="date">30/07/2010 </div>
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<div id="logbook">Genomic DNA of <em>Lactobacillus. Plantarum </em>WCFS1was extracted. Next, PCR of plnB from  the genomic DNA was conducted and products were confirmed by agarose gel  electrophoresis.</div>
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<div id="logbook">Genomic DNA of <em>Lactobacillus Plantarum </em>WCFS1 was extracted. Next, PCR of <em>plnB</em> from  the genomic DNA was conducted and products were confirmed by agarose gel  electrophoresis.</div>
</div>   
</div>   
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<div id="leftpart">
<div id="leftpart">
<div id="date">17/08/2010 </div>
<div id="date">17/08/2010 </div>
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<div id="logbook">Fusion PCR of agrC-plnB was  performed and products were confirmed by agarose gel electrophoresis.</div>
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<div id="logbook">Fusion PCR of <em>agrC-plnB</em> was  performed and products were confirmed by agarose gel electrophoresis.</div>
</div>   
</div>   
<div id="leftpart1">
<div id="leftpart1">
<div id="date">20/08/2010 </div>
<div id="date">20/08/2010 </div>
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<div id="logbook">Double-strand DNA of plnA  promoter (124bp) was obtained. By following the protocol of fusion PCR, annealing  of the forward primer (79bp) with the reverse primer (73bp) and the extension  were accomplished.</div>
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<div id="logbook">Double-stranded DNA of <em>plnA</em> promoter (124bp) was obtained. </div>
</div>   
</div>   
<div id="leftpart1">
<div id="leftpart1">
<div id="date">26/08/2010 </div>
<div id="date">26/08/2010 </div>
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<div id="logbook">agrC-plnB was ligated into  pBluescript KS (+). Ligation products were transferred into <em>E.coli </em>DH10B competent cells by heat shock  transformation. Ligation was confirmed by colony PCR and plasmid digestion  test.</div>
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<div id="logbook"><em>agrC-plnB</em> was ligated into  pBluescript KS (+). Ligation products were transferred into <em>E.coli </em>DH10B competent cells by heat shock  transformation. Ligation was confirmed by colony PCR and plasmid digestion  test.</div>
</div>  
</div>  
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<div id="leftpart1">
<div id="leftpart1">
<div id="date">12/09/2010 </div>
<div id="date">12/09/2010 </div>
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<div id="logbook">agrC was ligated into  pBluescript KS (+). Ligation products were transferred into <em>E.coli</em> DH10B competent cells by heat shock  transformation. Ligation was confirmed by colony PCR and plasmid digestion test.</div>
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<div id="logbook"><em>agrC</em> was ligated into  pBluescript KS (+). Ligation products were transferred into <em>E.coli</em> DH10B competent cells by heat shock  transformation. Ligation was confirmed by colony PCR and plasmid digestion test.</div>
</div>   
</div>   
   
   
<div id="leftpart">
<div id="leftpart">
<div id="date">20/09/2010 </div>
<div id="date">20/09/2010 </div>
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<div id="logbook">Performed three-way ligation of SP with plasmid pMG36e.</div>
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<div id="logbook">Performed three-way ligation of SP with plasmid pMG36e.</div>
</div>   
</div>   
<div id="leftpart">
<div id="leftpart">
<div id="date">22/09/2010 </div>
<div id="date">22/09/2010 </div>
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<div id="logbook">Colony PCR shown that the  insert was successfully ligated into plasmid pMG36e..</div>
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<div id="logbook">Colony PCR shown that the  insert was successfully ligated into plasmid pMG36e.</div>
</div>   
</div>   
<div id="leftpart1">
<div id="leftpart1">
<div id="date">24/09/2010 </div>
<div id="date">24/09/2010 </div>
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<div id="logbook"> gusA was ligated into  backbone pSB1C3. Ligation products were transferred into <em>E.coli </em>DH10B competent cells by heat shock transformation. Ligation  was confirmed by colony PCR and plasmid digestion test.</div>
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<div id="logbook"> <em>gusA</em> was ligated into  backbone pSB1C3. Ligation products were transferred into <em>E.coli </em>DH10B competent cells by heat shock transformation. Ligation  was confirmed by colony PCR and plasmid digestion test.</div>
</div>   
</div>   
<div id="leftpart">
<div id="leftpart">
<div id="date">26/09/2010 </div>
<div id="date">26/09/2010 </div>
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<div id="logbook">Sequencing of the insert was obtained  by DNA sequencing to further confirm the ligation product.</div>
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<div id="logbook">Sequencing of the insert (<em>gusA</em> inserted into pSB1C3) was obtained  by DNA sequencing to further confirm the ligation product.</div>
</div>   
</div>   
<div id="leftpart1">
<div id="leftpart1">
<div id="date">29/09/2010 </div>
<div id="date">29/09/2010 </div>
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<div id="logbook"> Two constructs agrC and agrC-plnB were transferred from pBluescript KS (+) to shuttle vector pMG36e. The ligation products were confirmed by colony PCR and enzyme digestion.</div>
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<div id="logbook"> Two constructs <em>agrC</em> and <em>agrC-plnB</em> were transferred from pBluescript KS (+) to shuttle vector pMG36e. The ligation products were confirmed by colony PCR and enzyme digestion.</div>
</div>   
</div>   
<div id="leftpart">
<div id="leftpart">
<div id="date">05/10/2010 </div>
<div id="date">05/10/2010 </div>
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<div id="logbook">Sequencing of four constructs: agrC in pBluescript KS (+), agrC-plnB in pBluescript KS (+), agrC in
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<div id="logbook">Sequencing of four constructs: <em>agrC</em> in pBluescript KS (+), <em>agrC-plnB</em> in pBluescript KS (+), <em>agrC</em> in pMG36e and <em>agrC-plnB</em> in pMG36e. All of them were finally confirmed.</div>
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pMG36e and agrC-plnB in pMG36e. All of them were finally confirmed.</div>
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</div>   
</div>   
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<div id="leftpart1">
<div id="leftpart1">
<div id="date">12/10/2010 </div>
<div id="date">12/10/2010 </div>
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<div id="logbook">GUS assay with the substrate  X-Gluc. E.coli containing PBI121 gave more gusA expression than control group.<br />
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<div id="logbook">GUS assay with the substrate  X-Gluc was performed. <em>E.coli</em> containing PBI121 gave more gusA expression than control group.<br />
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agrC-plnB (for localization test) was ligated into pBluescript SK (+). Ligation product was
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<em>agrC-plnB</em> (for localization test) was ligated into pBluescript SK (+). Ligation product was confirmed by colony PCR and enzyme digestion.
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confirmed by colony PCR and enzyme digestion.
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</div>
</div>
</div>   
</div>   
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<div id="leftpart">
<div id="leftpart">
<div id="date">13/10/2010 </div>
<div id="date">13/10/2010 </div>
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<div id="logbook">GUS assay with the substrate 4-NPG. E.coli containing PBI121 gave more gusA expression than control group.
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<div id="logbook">GUS assay with the substrate 4-NPG. <em>E.coli</em> containing PBI121 gave more <em>gusA</em> expression than control group.
</div>
</div>
</div>  
</div>  
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<div id="leftpart">
<div id="leftpart">
<div id="date">15/10/2010 </div>
<div id="date">15/10/2010 </div>
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<div id="logbook">Construct agrC-plnB-mCherry was built by ligating mCherry into pBluescript SK (+) containing
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<div id="logbook">Construct <em>agrC</em>-<em>plnB</em>-mCherry was built by ligating mCherry into pBluescript SK (+) containing
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agrC-plnB already.
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<em>agrC-plnB</em> already.
</div>
</div>
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</div>  

Latest revision as of 15:11, 26 October 2010

Team: HKUST

Lab Notebook

14/06/2010
BioBrick BBa_I746101 was extracted from 96 wells plates. E.coli DH10B competent cells were successfully transformed with BioBrick BBa_I746101 which contains agrC.
25/06/2010
Plasmid PMH4 containing mCherry and plasmid PBI121 conatining gusA reporter gene were successfully transferred into E.coli DH10B competent cells.
12/07/2010
Fusion PCR of agrC-mCherry was performed and products were confirmed by agarose gel electrophoresis.
21/07/2010
Successfully extract plasmid pMG36e out from filter paper.
22/07/2010
Transform E.coli DH10B competent with plasmid pMG36e.
28/07/2010
Plasmid pMG36e was enzyme digested and confirmed..
30/07/2010
Genomic DNA of Lactobacillus Plantarum WCFS1 was extracted. Next, PCR of plnB from the genomic DNA was conducted and products were confirmed by agarose gel electrophoresis.
10/08/2010
Midi-prep of shuttle vector pMG36e was performed and extracted plasmids were confirmed by enzyme digestion test.
16/08/2010
PCR of gusA from PBI121 was conducted and products were confirmed by agarose gel electrophoresis.
17/08/2010
Fusion PCR of agrC-plnB was performed and products were confirmed by agarose gel electrophoresis.
20/08/2010
Double-stranded DNA of plnA promoter (124bp) was obtained.
26/08/2010
agrC-plnB was ligated into pBluescript KS (+). Ligation products were transferred into E.coli DH10B competent cells by heat shock transformation. Ligation was confirmed by colony PCR and plasmid digestion test.
28/08/2010
Primers arrived and prepareed primers for ligation. Double strand DNA of FLAG-tag, both part 1&2 of signal peptide and DD13-RIP was obtained.
12/09/2010
agrC was ligated into pBluescript KS (+). Ligation products were transferred into E.coli DH10B competent cells by heat shock transformation. Ligation was confirmed by colony PCR and plasmid digestion test.
20/09/2010
Performed three-way ligation of SP with plasmid pMG36e.
22/09/2010
Colony PCR shown that the insert was successfully ligated into plasmid pMG36e.
24/09/2010
gusA was ligated into backbone pSB1C3. Ligation products were transferred into E.coli DH10B competent cells by heat shock transformation. Ligation was confirmed by colony PCR and plasmid digestion test.
26/09/2010
Sequencing of the insert (gusA inserted into pSB1C3) was obtained by DNA sequencing to further confirm the ligation product.
29/09/2010
Two constructs agrC and agrC-plnB were transferred from pBluescript KS (+) to shuttle vector pMG36e. The ligation products were confirmed by colony PCR and enzyme digestion.
05/10/2010
Sequencing of four constructs: agrC in pBluescript KS (+), agrC-plnB in pBluescript KS (+), agrC in pMG36e and agrC-plnB in pMG36e. All of them were finally confirmed.
11/10/2010
mCherry was ligated into pBluescript SK (+). Some colonies on the plate showed red color expressed by mCherry.
12/10/2010
GUS assay with the substrate X-Gluc was performed. E.coli containing PBI121 gave more gusA expression than control group.
agrC-plnB (for localization test) was ligated into pBluescript SK (+). Ligation product was confirmed by colony PCR and enzyme digestion.
13/10/2010
GUS assay with the substrate 4-NPG. E.coli containing PBI121 gave more gusA expression than control group.
15/10/2010
Construct agrC-plnB-mCherry was built by ligating mCherry into pBluescript SK (+) containing agrC-plnB already.